title: ESCRT requirements for EIAV budding.
abstract: BACKGROUND
  Retroviruses and many other enveloped viruses usurp the cellular ESCRT pathway to bud from cells. However, the stepwise process of ESCRT-mediated virus budding can be challenging to analyze in retroviruses like HIV-1 that recruit multiple different ESCRT factors to initiate budding.
  
  
  RESULTS
  In this study, we characterized the ESCRT factor requirements for budding of Equine Infectious Anemia Virus (EIAV), whose only known direct ESCRT protein interaction is with ALIX. siRNA depletion of endogenous ESCRT proteins and "rescue" experiments with exogenous siRNA-resistant wild type and mutant constructs revealed budding requirements for the following ESCRT proteins: ALIX, CHMP4B, CHMP2A and VPS4A or VPS4B. EIAV budding was inhibited by point mutations that abrogate the direct interactions between ALIX:CHMP4B, CHMP4B:CHMP2A, and CHMP2A:VPS4A/B, indicating that each of these interactions is required for EIAV budding. Unexpectedly, CHMP4B depletion led to formation of multi-lobed and long tubular EIAV virions.
  
  
  CONCLUSIONS
  We conclude that EIAV budding requires an ESCRT protein network that comprises EIAV Gag-ALIX-CHMP4B-CHMP2A-VPS4 interactions. Our experiments also suggest that CHMP4B recruitment/polymerization helps control Gag polymerization and/or processing to ensure that ESCRT factor assembly and membrane fission occur at the proper stage of virion assembly. These studies help establish EIAV as a streamlined model system for dissecting the stepwise processes of lentivirus assembly and ESCRT-mediated budding.
authors.affiliation: Department of Biochemistry, University of Utah School of Medicine, Salt Lake City 84112-5650, Utah, USA. wes@biochem.utah.edu.
authors.forename: Virginie
authors.initials: V
authors.lastname: Sandrin
authors.forename: Wesley I
authors.initials: WI
authors.lastname: Sundquist
mesh_terms: D002460:Cell Line; D056827:Endosomal Sorting Complexes Required for Transport / Q000378:metabolism*; D054884:Host-Pathogen Interactions; D006801:Humans; D004860:Infectious Anemia Virus, Equine / Q000502:physiology*; D011485:Protein Binding; D025941:Protein Interaction Mapping; D057074:Virus Release
chemical_list: D056827:Endosomal Sorting Complexes Required for Transport